Peanut components measured by ISAC: comparison with ImmunoCap and clinical relevance in peanut allergic children

Background Specific IgE (sIgE) against the peanut component Arachis hypogaea (Ara h) 2 has been shown to be the most important allergen to discriminate between peanut allergy and peanut tolerance. Several studies determined sIgE cut off values for Ara h 2, determined by singleplex measurements. However, cut off values for Ara h 2 from multiplex arrays are less well defined. The aim of this study was to evaluate the correlation between Ara h 2 sIgE determined by singleplex versus multiplex measurements and to assess the diagnostic value of the different peanut components included in Immuno Solid-phase Allergen Chip (ISAC) multiplex analysis in children with a suspected peanut allergy. Methods In this retrospective study we analyzed Ara h 2 sIgE values with singleplex Fluorescence Enzyme Immunoassay (FEIA, ImmunoCap) and multiplex microarray (ISAC) measurements in 117 children with a suspected peanut allergy. Also, other peanut components measured by ISAC were analyzed. Double blinded placebo controlled oral food challenges were used as golden standard. Results Among all studied peanut components FEIA Ara h 2 sIgE showed the highest area under the curve (AUC, 0.922), followed by ISAC Ara h 6 and Ara h 2 sIgE with AUCs of respectively 0.906 and 0.902. Best cut off values to diagnose peanut allergy were 4.40 kU/l for FEIA Ara h 2 sIgE and, 7.43 ISU and 8.13 ISU for respectively Ara h 2 and Ara h 6 sIgE in ISAC microarray. Ara h 2 sIgE determined in FEIA and ISAC showed a good correlation (r = 0.88; p < 0.01). Conclusion Ara h 6 and Ara h 2 sIgE in multiplex ISAC are both good predictors of clinical peanut allergy in Dutch children, and their performance is comparable to the use of Ara h 2 in singleplex FEIA. The simultaneous measurement of different peanut components using ISAC is an advantage and clinically useful to detect peanut allergic children that are Ara h 2 negative but sensitized to other peanut proteins such as Ara h 6.

Electrochemical Immunosensor for the Simultaneous Determination of Two Main Peanut Allergenic Proteins (Ara h 1 and Ara h 6) in Food Matrices

Efficiently detecting peanut traces in food products can prevent severe allergic reactions and serious health implications. This work presents the development of an electrochemical dual immunosensor for the simultaneous analysis of two major peanut allergens, Ara h 1 and Ara h 6, in food matrices. A sandwich immunoassay was performed on a dual working screen-printed carbon electrode using monoclonal antibodies. The antibody–antigen interaction was detected by linear sweep voltammetry through the oxidation of enzymatically deposited silver, which was formed by using detection antibodies labeled with alkaline phosphatase and a 3-indoxyl phosphate/silver nitrate mixture as the enzymatic substrate. The assay time was 2 h 20 min, with a hands-on time of 30 min, and precise results and low limits of detection were obtained (Ara h 1: 5.2 ng·mL−1; Ara h 6: 0.017 ng·mL−1). The selectivity of the method was confirmed through the analysis of other food allergens and ingredients (e.g., hazelnut, soybean and lupin). The dual sensor was successfully applied to the analysis of several food products and was able to quantify the presence of peanuts down to 0.05% (w/w). The accuracy of the results was confirmed through recovery studies and by comparison with an enzyme-linked immunosorbent assay. Tracking food allergens is of utmost importance and can be performed using the present biosensor in a suitable and practical way.

Peanut Components Measured by ISAC: Comparison With ImmunoCap and Clinical Relevance in Peanut Allergic Children

BackgroundSpecific IgE (sIgE) against the peanut component Arachis hypogaea (Ara h) 2 has been shown to be the most important allergen to discriminate between peanut allergy and peanut tolerance. Several studies determined sIgE cut off values for Ara h 2, determined by singleplex measurements. However, cut off values for Ara h 2 from multiplex arrays are less well defined. The aim of this study was to evaluate the correlation between Ara h 2 sIgE determined by singleplex versus multiplex measurements and to assess the diagnostic value of the different peanut components included in Immuno Solid-phase Allergen Chip (ISAC) multiplex analysis in children with a suspected peanut allergy.Methods In this retrospective study we analyzed Ara h 2 sIgE values with singleplex Fluorescence Enzyme Immunoassay (FEIA, ImmunoCap) and multiplex microarray (ISAC) measurements in 117 children with a suspected peanut allergy. Also, other peanut components measured by ISAC were analyzed. Double blinded placebo controlled oral food challenges were used as golden standard.Results Among all studied peanut components FEIA Ara h 2 sIgE showed the highest area under the curve (AUC, 0.922), followed by ISAC Ara h 6 and Ara h 2 sIgE with AUCs of respectively 0,906 and 0,902. Best cut off values to diagnose peanut allergy were 4.40 kU/l for FEIA Ara h 2 sIgE and, 7.43 ISU and 8.13 ISU for respectively Ara h 2 and Ara h 6 sIgE in ISAC microarray. Ara h 2 sIgE determined in FEIA and ISAC showed a good correlation (r=0.88. p<0.01). ConclusionAra h 6 and Ara h 2 sIgE in multiplex ISAC are both good predictors of clinical peanut allergy, and their performance is comparable to the use of Ara h 2 in singleplex FEIA. The simultaneous measurement of Ara h 6 and Ara h 2 sIgE using ISAC is an advantage and clinically useful to detect peanut allergic children that are monosensitized to Ara h 6.

Oxidative Stability of Protease Treated Peanut with Reduced Allergenicity

Oxidative stability and allergenicity are two major concerns of peanuts. This study evaluated the impact of protease treatment of peanuts on its oxidative stability during storage. The raw and dry-roasted peanut kernels were hydrolyzed with Alcalase solution at pH 7.5 for 3 h. The contents of Ara h 1, Ara h 2, and Ara h 6 in peanuts were determined before and after enzyme treatment by a sandwich ELISA. After drying, the samples were packed in eight amber glass jars and stored at 37 °C for 1–8 weeks. Controls are untreated raw and dry-roasted peanuts packed and stored in the same way as their treated counterparts. Samples were taken biweekly to determine peroxide value (PV) and thiobarbituric acid reactive substances (TBARS) as indicators of oxidation (n = 3), and to determine antioxidant activity. Alcalase treatment reduced intact major allergens Ara h 1, Ara h 2, and Ara h 6 by 100%, 99.8%, and 85%, respectively. The PVs of Alcalase-treated raw and roasted peanuts was lower than those of untreated (p < 0.05) over the 8-week storage. The TBARS of Alcalase-treated raw peanuts were slightly higher than that of untreated (p < 0.05), but the TBARS of Alcalase-treated dry-roasted peanuts were slightly but significantly lower than that of untreated (p < 0.05). The protease treatment increased the antioxidant activities including reducing power, DPPH free radical scavenging capacity, and metal chelating capacity of peanuts.